COLLEGE'S NAME LABUAN MATRICULATION COLLEGE

STUDENT'S NAME MOHAMMAD ADAM BIN ABDULSAN

MATRICULATION NUMBER MS2415114975

PRACTICUM GROUP H40

TITLE BASIC TECHNIQUES OF MICROSCOPY

DATE 15 JULY 2024

LECTURER'S NAME MOHD BADRUDDIN BIN ABDULLAH

EXPERIMENT 1: BASIC TECHNIQUES OF MICROSCOPY

 Objective(s):
1. Obtain accurate images.
2. Determine the depth of field.
3. Determine the field of view.
4. Calculate the acctual magnification.
5. Apply the use of oil immersion with high magnification (oil immersion only)

(1.1) Images, Depth of Fields and Field of View of the Microscope.

(1.1.1) Images under the microscope

 Procedure:
1. The 'e' prepared slide was observed using 4x objective lens.

 Observation:
'e' prepared slide

e
Actual magnification: 40x
 (1.1.2) The depth of field

 Procedure:
1. The position of the thread on the slide was observe with naked eyes and
the colour of the thread had been identified.
2. The cross threads under the microscope was observed using 4x and 10x
objective lens.

 Observation:

Cross thread prepared slide

Bottom: Yellow

Middle: Red

Top: Blue

Actual magnification: 40x

 (1.1.3) The field of view.

 Procedure:
1. The transparent ruler was placed on the stage. Make sure the ruler scale is
placed at the centre of the field of view
2. The transparent ruler was observed using the 4x, 10x and 40x objective
lenses. (Increase the amount of light by adjusting the control knob to the
maximum).
3.
4. The diameter of field of view had been determined at 4x, 10x and 40x
objective lens.
5. The diameter of field of view had beeb calculated using formula below.

Diameter of field of view under

low magnification power high magnification power
---------------------------------------------------- = -----------------------------------
Diameter of field of view under low magnification power
high magnification power

 Observation:
Table 1.1 Diameter field of view

Objective lens Diameter in mm Diameter in um

4x 3.0 14.2
10x 2.0 11.4
40x 1.0 8.7
Ruler scale prepared slide.

Actual magnification: 40x

(1.2) Magnification.

 Procedure:
1. The actual magnification of a specimen had been determined using the
formula below.

Actual Magnification power of Magnification power of

= X
magnification objective lens ocular lens

2. The actual magnification had been calculated.

 Observation:
Table 1.2 Actual magnification of a specimen

Actual magnification
Magnification Magnification power of objective lens
power of
4x 10x 40x 100x
ocular lens
10x 40x 100x 400x 1000x
(1.3) Oil Immersion Objective Lens.

 Procedure:
1. The prepared slide under the microscope observed using 100x objective
lens.

 Observation:

Paramecium sp. prepared slide.

Actual magnification: 40x

 Discussion:
1. Throughout this experiment, the specimens and objects were
observed using a light compound microscope under 4X, 10X, 40X
and 100X magnification with the use of immersion oils.

2. When the 'e' prepared slide was observed under the microscope it
was in an inversed position. This is because of the focal length of the
objective lens. The image focused by the lens crosses over before the
eyepiece further magnifies the image of the letter 'e'. Then the
objective lens inverts the image because of the lenses curvature. This
causes the image of the letter 'e' to be inverted at the focal length.

3. When the cross threads slide was observed under 4X and10X

magnification, the arrangement of the threads remained unchanged.
This is because depth of field is inversely proportional to
magnification. This means, that when the magnification of the
objective lens increases, the depth of field decreases. Therefore, the
arrangement of the cross threads is the same.

4. When the slide of the transparent ruler was observed under the
microscope, the field of view became smaller as the magnification
increased. This is because, field of view in inversely proportional to the
magnification of the objective lens. The transparent ruler appears larger
when the magnification is higher because only a small area of the ruler
is spread out to cover the field of view through the eyepiece.

5. After using the formula to calculate actual magnification, it was

determined that when the magnification power of ocular lens and
objective lens increases, then the actual magnification increases. This
is because the standard magnification of ocular lens is 10X. When it is
multiplied to the magnification of the objective lens which is 4X, 10X,
40X and 100X it provides the value of actual magnification which is
higher .

6. When observing the gram stain mix specimen under the

microscope, the oil immersion lens was used. Before observing the
specimen, a few drops of immersion oil were placed on the cover slip
over that area, and very carefully the oil immersion lens was put into
place.
7. The microscope magnifies the image of the specimen by bending
light. When the magnification is higher, the light is bent more. At a
certain point, the light is bent so much that it can't make it through
the objective lens. At that point which is around 100X, a few drops of
immersion oil should be placed between the specimen and the
objective lens. The oil "unbends" the light to stretch out the working
distance and make it possible to observe the image of the specimen
at high magnifications.

8. Immersion oil is also important to improve the power of resolution.

This is because when light travels from a material of one refractive
index to material of another, it bends. Light at different wavelengths
bends at different angle and causes the magnified images to be less
distinct. Oil immersion eliminates two refractive surfaces, so that the
magnifications of 100x can be achieved while still preserving good
resolution. Due to this, a clear image of the Paramecium sp. can be drawn.

9. After the specimen was observed and drawn, the oil immersion
objective lens was cleaned by using lens paper. This is an important
step to avoid the immersion oil from smudging and damaging the lens.

 Conclusion:
As stated in the hypothesis before, the image produced by the
microscope is inverted from its original position which was tested
through the observation of the 'e' prepared slide.
Secondly, when the magnification increases, the depth of field decreases,
which was tested through the observation of a prepared cross thread slide.
Third, when the magnification increases, the field of view becomes
smaller which was tested by observing a prepared transparent ruler
slide.
Next, when the magnification of objective lens and ocular lens
increases, then the actual magnification increases which was tested
by the usage of the formula.
Lastly, the use of the oil immersion objective lens with high magnification
produces a high-resolution image of the specimen which was tested by
observing a prepared gram stain mix slide.
 Questions:

 Part A
1. Which is the shortest objective lens?
= Scanning objective lens.
2. Which objective lens should you use when you begin to focus a spesimen?
= Scanning objective lens.
3. Which objective lens should be in positions before you store a
microscope?
= Scanning objective lens.
4. Which objective lens will deliver the highest amount of light?
= High power objective lens.
5. Which objective lens requires immersion oil to fill up the space between
the specimen and the lens?
= Oil immersion objective lens.

6. Which objective lens will still remain in focus when place at the longest
working distance from the specimen?
= Scanning objective lens.
7. When using an ocular lens with 10x magnification power, which
objective lens should be used to obtain the following actual
magnification?
a. 100 times of its diameter
= Low power objective lens.
b. 1000 times of its diameter
= High power objective lens.
 Part B
1. Using 40x objective lens, determine the length and width of a cell from
a piece of cork tissue with approximately 20 cells in horizontal position
and 10 cells in vertical position.

Figure 1.4 Cork tissue

Actual magnification = (40 x 10)
= 400x
Considering the diameter of objective lens is 30 mm,
Horizontal position, 30 / 20 = 1.5 mm
Vertical position, 30 / 10 = 3.0 mm
Hence, Length, 1.5 / 400 = 0.00375 mm = 3.75 um
Width, 3.0 / 400 = 0.0075 mm = 7.5 um
2. Based on laboratory practices, what do you use to clean the microscope
lenses?
= Lens paper
3. While observing a moving microorganism under a microscope, you
found that the organism has moved out of the field of view to the right.
In order to keep observing the microorganism, which direction do you
move your slide (right/left)?
= To the left
4. How do you adjust the slide when the specimen is out of the field of
view to the top?
= Adjust the slide downwards carefully and slowly to avoid losing sight of the
specimen or damaging the microscope lens.
 Part C
1. Complete the following sentences.
a. A microscope is called a compound microscope when it consists of
more than one set of objective and ocular lens.
b. Condenser and iris diaphragm are useful to coordinate
and control light intensity.
 Reference:

Matriculation Division, Ministry of Education Malaysia.(2022).Biology

Laboratory Manual, Semester I & II, SB015 & SB025.(Thirteenth Edition).
Matriculation Division Ministry of Education Malaysia.

Studocu. (30 July 2021). LAB Report EXP 1 (SEM 1) ( Marked).

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